HIV-specific cytotoxic T lymphocyte (CTL) responses have been implicated in viral control, but precise correlates of immune protection remain to be defined. One parameter likely to be associated with viral control is the potential of CTLs to cope with the extensive viral sequence variation. The ability of HIV- specific CTLs to recognize epitope variants arising in vivo as potential escape mutants has been studied for a few, highly selected epitopes. However, a more systematic approach of assessing the capacity of CTLs to recognize frequently occurring sequence variants (i.e. potentially infecting sequences) has not been undertaken. Similarly, the mechanisms involved in broad recognition of epitope variants have not completely been assessed. The goal of this proposal is to determine the potential of HIV Gag-specific T cells to recognize frequently occurring sequence variants, and to link variant recognition with the functional avidity of these responses. To this end, immune responses in subjects with and without spontaneous control of viral replication will be tested using 11mer Gag peptides overlapping by 10 amino acids, including all variant 11mer sequences that are present in at least 5% of all Gag sequences available at the Los Alamos National Laboratory Sequence database. The functional avidity of CTLs specific for all targeted epitopes and their variants will be determined by assessing the peptide concentration needed to elicit half-maximal responses in IFN-3/IL-2 dual color ELISpot assays, and correlated with disease progression as well as the CTLs'ability to recognize large proportions of naturally occurring variants. Furthermore, the functional avidity of Gag-specific CTLs will be determined in subjects on antiretroviral treatment prior to treatment interruptions and associated to the viral load set-point after cessation of treatment to test if high-avidity responses are predictive of superior outcome of treatment interruptions compared to low-avidity responses. In summary, this application will provide critical data correlating the functional avidity of Gag-specific CTL with outcome of HIV disease and offering a potential mechanistic explanation for this benefit through the increased recognition of viral variants. These data will be vital for immunogen design, since a broadly applicable prophylactic vaccine will need to cope effectively with viral sequence variability. PUBLIC HEALTH RELEVANCE: The present application aims to define correlates of immune protection to HIV infection by assessing two poorly defined functions of HIV-specific T cells. Due to the extreme variability of HIV, the ability of T cells to cope with HIV antigenic diversity as well as their affinity for even low concentrations of HIV antigen are important parameters to be considered in vaccine design. In this proposal, both of these functions will be assessed in subjects with and without spontaneous control of HIV replication and in individuals undergoing antiretroviral treatment interruptions to address the role of highly avid HIV- specific T cells in the immune control of this pathogen.